Over the past three decades, dietary restriction (DR) has become the gold standard against which manipulations that retard aging are compared. Because DR has been shown to increase the lifespan of a wide variety of organisms ranging from invertebrates to rodents, DR is viewed as a universal aging intervention. However, a recent study suggests that the genotype of an animal is a major determinant in the ability of the animal to respond to DR, e.g., two-thirds of the 41 recombinant inbred (RI) lines of mice studied either did not respond or showed reduced lifespan when fed 40% DR (40% less diet than that consumed by mice fed ad libitum). The overall goal of this grant is to explore the interaction between genotype and the level of DR using nine genetically diverse RI lines of mice. We hypothesize that DR will increase lifespan and delay aging in all genotypes; however, the effect of DR will be both genotype- and dose-dependent, i.e., one level of DR is not optimal for all genotypes. The comprehensive design of this study, allows us for the first time to obtain an accurate view of how genetic diversity impacts the effect of DR on lifespan and aging. Specific Aim 1: To determine whether DR affects lifespan in a genotype- and dose-dependent manner. The lifespans of nine RI lines of female mice that show maximum diversity in response to 40% DR will be fed ad libitum or 10, 20, or 40% less diet than the amount consumed by each RI line fed ad libitum. Based on our hypothesis, we predict that all nine RI lines will show an increase in lifespan; however, the level of restriction necessary to obtain an increase in lifespan will vary with genotype. Specific Aim 2: To determine whether DR affects end-of-life pathology in a genotype- and dose-dependent manner. Because a reduction in most age-related pathologies is a hallmark feature of DR, we will conduct a comprehensive pathological analysis of all the female mice that die in Aim 1. We predict that the increase in lifespan by DR will be accompanied by a reduction/delay in most age-related pathological lesions, which would support the premise that DR increases lifespan by retarding aging. We also predict that the effect of DR on end-of-life pathological lesions will be genotype- and DR dose-dependent. Specific Aim 3: To identify potential pathways involved in the anti-aging mechanism of DR. We will measure the levels of transcripts in RNA isolated from liver and epididymal fat of mice fed AL and the three DR diets for 12 months using Illumina Mouse microarrays. Employing unbiased analyses of our microarray data, we will identify pathways that are altered significantly by DR. Because we will have transcriptome data on 27 cohorts of mice fed DR (nine strains of mice fed 10, 20, and 40% DR), we are in a unique position to identify gene profiles that have the greatest predictive power to identify the cohorts of mice that show a significant increase in lifespan when placed on a DR diet, and from these data, we can identify potential mechanisms key in the life-extending action DR.